In the Breastfeeding, Antiretrovirals and Nutrition Study (BAN), 28 weeks of maternal highly active antiretroviral therapy (HAART) started at delivery decreased transmission by 52% and 28 weeks of extended infant NVP decreased transmission by 74% over the control arm (sdNVP and 7 days of Combivir). Since adherence, as measured by pill counts or self report, does not capture bioavailability or efficacy of the antiretrovirals, we will measure drug concentrations and plasma and breastmilk viral load (VL) and correlate these data with transmission. Increases in breastmilk HIV VL likely play an important role in breast milk transmission. Possible causes of increases in VL include breast problems (e.g. mastitis or cracked nipples), febrile illnesses, non-adherence to the treatment regimens, or poor bioavailability of the antiretrovirals. Plasma levels of HIV RNA and breastmilk HIV RNA and DNA will be measured and correlated with mastitis, and/or maternal fevers (Specific Aim 1). Drug concentrations in maternal plasma and breastmilk, and infant plasma will be measured (Specific Aim 2), and adherence to study drugs will be determined (Specific Aim 3). Viral loads, drug levels and adherence will be correlated with transmission to the infant. The BAN study did not have an arm that evaluated the combined regimen of both maternal HAART plus extended infant NVP, but data generated in Aims 1-3 will be used to model what this effect might have been (Specific Aim 4). With samples at more frequent time points than most clinical trials, results from this work will be able to determine potential causes of increased breastmilk viral load in women receiving HAART. To understand differential drug penetration more fully, concentration-effect relationships need to be generated. No study has published associations between adherence rates of maternal ART during breastfeeding and the breastmilk concentrations, and ultimately concentrations in the infant plasma;nor has any study evaluated infant prophylaxis adherence based on bottle weights. No data exist to directly estimate the efficacy of combining maternal ART with infant prophylaxis for prevention of breastmilk transmission. In short, no study has used the comprehensive approach of determining associations of measured adherence, antiretroviral concentrations, and VL with breastmilk transmission which this proposal aims to do. Relevance: The data collected through this comprehensive approach will be crucial for the development of strategies that will prevent future breastmilk transmissions, for instance, by improving adherence, or using combination maternal HAART and infant prophylaxis. PUBLIC HEALTH RELEVANCE: The Breastfeeding, Antiretrovirals and Nutrition (BAN) Study enrolled 2370 HIV-infected women and their infants to examine different ways to prevent breast milk transmission of HIV. Despite the interventions, some transmissions occurred. We will measure the levels of HIV RNA, HIV DNA, and antiretroviral drug concentrations in plasma and breast milk, as well as determine adherence to study drug regimens in participants from this study to determine why each case of breast milk transmission occurred. We will also review data on cases of maternal fever or breast problems (mastitis, cracked nipples, etc) and correlate these with increases in breast milk viral load and HIV transmission. We will model the potential effect of using combination infant prophylaxis with maternal HAART for the prevention of breast milk transmission of HIV. With these data we will be able to develop strategies that will prevent future transmissions, such as improving adherence, or using combination maternal HAART and infant prophylaxis.